Radiation therapy is an important treatment modality for cancer patients and is used for treating a wide variety of different tumor types. Most (more than 70%) of the DNA damages produced by ionizing radiation during treatment are those repaired by the base excision repair system and ablation of base excision repair greatly increases cellular radiation sensitivity in model systems. Both low and higher LET radiation used in the treatment of a variety of cancers produces clustered DNA lesions which include the many damages recognized and removed during base excision repair of single lesions. During the prior funding period, we established that attempted but abortive repair of radiation-induced clustered DNA damages leads to an increase in the number of potentially lethal double strand breaks and an increase in cellular lethality. We also showed that this increase in the formation of double strand breaks and cellular lethality was mediated solely by the DNA glycosylases that recognize the initial base lesions. Thus, manipulation of the substrate specificities of the DNA glycosylases that recognize radiation damage could be used to potentiate DNA damage during radiotherapy and significantly influence treatment outcome and therapeutic gain. The overall goal of the proposed research is to delineate the substrate specificities of the DNA glycosylases that initiate base excision repair of radiation damage and as well produce potentially lethal double strand breaks at radiation-induced clustered lesions. The ultimate goal is to develop strategies based on structure/function relationships that would increase the effective dose to the tumor without damaging the normal tissue.